A. Carpentieri et al. / Pharmacological Research 65 (2012) 437–444Fig. 1. Metabolic pathway of melatonin synthesis.
direct antioxidant, as a chemotoxicity reducing agent and a putative
cerebrospinal ﬂuid and reaches other body ﬂuids such as bile, cere-
anti-aging substance
brospinal ﬂuid, saliva, semen, ovarian follicular ﬂuid and amniotic
ﬂuid. Small amounts of unmetabolized MEL are excreted in the
1. Structure, metabolism and secretion
The half life of MEL in the serum has been calculated to be
MEL is an indoleamine containing two functional groups, which
in the range 30–50 min The ability to synthesize MEL is
are decisive not only for the receptor binding but also for its
rather constant in a given person, but there is marked variability
amphiphilicity giving to the molecule the capacity to enter any cell
among individuals. There is evidence that MEL levels decrease with
compartment or body ﬂuid. Due to its extensive solubility in lipids,
increasing age in mammals, including humans. The serum level of
MEL easily passes by diffusion from the peripheral circulation to
MEL is very low in the ﬁrst weeks of postnatal life, without diur-
other ﬂuids or cells. In serum, 70% of the MEL is bound to albumins
nal variation. At six months of life the typical diurnal rhythm of
and the remaining 30% diffuses to the surrounding tissues
secretion appears, reaching the maximum levels between the third
Tryptophan and serotonin are precursors of MEL. Two well char-
and sixth years of life. A marked decrease in MEL secretion has
acterized enzymes participate in its synthesis: N-acetyltransferase
been noted during sexual maturation. At 40–50 years, a noticeable
(AANAT), which converts serotonin to N-acetylserotonin (NAS), and
decrease in daily MEL synthesis has been observed and after the 70
hydroxyindole-O-methyltransferase (HIOMT), which converts NAS
years of age the diurnal rhythm of secretion practically disappears
to MEL (The regulation of MEL synthesis is controlled by
in most individuals seasonal variation in the synthesis of
the light–dark cycle, acting through neural activation of the ante-
MEL in humans seems to exist, the levels being higher in winter
rior hypothalamus via the axons of retinal ganglion cells running
in the optic nerves and forming the retino-hypothalamic tract.
The catabolism of MEL was thought to be almost exclusively
Suprachiasmatic nucleus (SCN) is connected with the pineal gland
done by the hepatic P450 monooxygenases, followed by conju-
through paraventricular nuclei and preganglionic sympathetic neu-
gation of the resulting 6-hydroxy-melatonin to give the main
rons. Norepinephrine released from postganglionic sympathetic
urinary metabolite 6-sulfatoxy-melatonin. This might occur with
ﬁbers at pinealocyte membrane stimulates its adrenoceptors lead-
the circulating hormone. In contrast, in the central nervous sys-
ing to cAMP formation as well as other second messengers, which
tem (CNS) the oxidative pyrrole-ring cleavage predominates and
stimulate the expression and activity of AANAT, the ﬁrst-rate limit-
no 6-hydroxy-melatonin was detected after MEL injection into the
ing enzyme in MEL production non-mammalian vertebrates,
cisterna magna, which may be important because much more MEL
this enzyme appears to be directly controlled by circadian clock
is released via the pineal recess into the cerebrospinal ﬂuid than
genes in the pineal gland
into the circulation. The primary cleavage product is N1-acetyl-N2-
The secretion of MEL has a typical diurnal rhythm. At night
formyl-5-methoxykynuramine (AFMK). Several different reactions
the synthesis and secretion of MEL are stimulated, reaching a
lead to the same product, AFMK, and this pathway contributes to
peak value (80–150 pg/mL) between midnight and 3 a.m., while its
about one third of the total catabolism. AFMK is converted into
concentration during the day is low (10–20 pg/mL) syn-
N1-acetyl-5-methoxykynuramine (AMK). AFMK and AMK form
thesized in the pineal gland, MEL is secreted into the blood and
metabolites by interaction with reactive oxygen and nitrogen
A. Carpentieri et al. / Pharmacological Research 65 (2012) 437–444
species. Some other metabolites have been detected, but appar-
4. MEL and mitochondria
ently in minor quantities
The lipophilic nature of MEL favors that the indolamine crosses
cell membranes to easily reach subcellular compartments includ-
2. MEL receptors
ing mitochondria. MEL interacts with lipid bilayers and stabilizes
mitochondrial inner membranes, which may improve the elec-
Most actions of MEL are mediated by membrane receptors and
tron transport chain (ETC) activity. At a concentration of 1 nM,
nuclear sites corresponding to orphan members of the nuclear
MEL increases the activity of the complexes I and IV in rat liver
receptor superfamily RZR/ROR. Three subtypes of mammalian MEL
mitochondria, while 10–100 nM MEL stimulates the activity of
receptors have been proposed and cloned. Two of these, MT1 and
those complexes in brain mitochondria. When a dose of cyanide
MT2, are members of the 7-transmembrane G-protein-coupled
decreases the complex IV activity by 50%, MEL counteracts this inhi-
receptor (GPCR) family. These two receptors are classiﬁed as unique
bition. In contrast, when the complex IV is totally inactivated by
subtypes based on their molecular structure and chromosomal
cyanide, MEL is unable to counteract this inhibition, independently
localization. Both of them belong to the class A group of rhodopsin-
of its concentration. The previous data suggest that the regulation
like GPCRs. They are formed by 350 and 362 aminoacids and their
by MEL on the activities of complexes I and IV does not only rely
calculated molecular weights are 39 and 40 kDa, respectively. The
on its antioxidant properties. MEL has a high reduction potential,
gene MTRN1A for the MT1 receptor is located at position 4q35-
which suggests that MEL might interact with the components of
1 and the gene MTNR1B for the MT2 receptor at 11q21-22. They
the electron transport chain increasing the electron ﬂow, and con-
share 60% homology. MT1 has two potential glycosylation sites in
sequently, the ATP production
the N-terminal region and MT2 has one potential glycosylation site
As known, reactive oxygen species (ROS) and reactive nitrogen
in the same region. Both receptors involve signaling through inhi-
species (RNS) are synthesized as subproducts of the mitochon-
bition of cAMP formation and protein kinase A activity, and effects
drial electron transport chain, although the main sources of •NO
on phospholipase A2 and C, calcium and potassium channels
are the reactions catalyzed by nNOS (neuronal nitric oxide syn-
MT3, the third receptor, is an enzyme identiﬁed as quinone reduc-
thase) and the iNOS (inducible nitric oxide synthase). Moderate
tase 2 (QR2). Little information exists on nuclear MEL receptors.
levels of •NO are considered favorable for mitochondrial function,
Another MEL-related receptor, named GPR50, has also been found
whereas high •NO concentrations produce severe ETC dysfunction,
in different species including humans
1c, the ﬁrst type of
blocking the respiration in extreme conditions of inﬂammation
MEL receptor discovered, is a receptor subtype expressed in non
seems to protect proteins of the ETC and mtDNA from
mammalian species is some evidence that MEL receptor
ROS/RNS-induced oxidative damage. The hormone limits the loss
expression exhibits circadian variation
of intramitochondrial GSH, improves the electron transport chain
activity and reduces mtDNA damage. It apparently increases the
3. MEL and antioxidant properties
expression and activity of complex IV and the activity of complex I,
which improves mitochondrial respiration and increases ATP syn-
There is a large body of evidence that MEL is a major scav-
enger of both oxygen and nitrogen-based reactive molecules. MEL
MEL also regulates the GSH redox status in isolated brain and
provokes this effect at both physiological and pharmacological con-
liver mitochondria when it is disrupted by oxidative stress. The
centrations. Several of its metabolites can also detoxify free radicals
indolamine increases the mitochondrial GSH content, decreases
and derivatives The indolamine eliminates the decomposi-
oxidized GSH (GSSG) and hydroperoxide levels and stimulates the
tion products of peroxynitrites, including free hydroxyl radicals and
activity of GPx and GSH reductase (GR), which are two enzymes
nitrogen dioxide radicals and the carbonate radical in the presence
involved in the GSH-GSSG balance et al.
of physiological CO2 concentration induces the synthesis
demonstrated that micromolecular concentrations of MEL prevent
of another intracellular antioxidant, glutathione (GSH), in rabbits
calcium dependent cardiolipin peroxidation in mitochondria, thus
that exhibit diabetes-induced oxidative stress beneﬁt of
protecting against the induction of mitochondrial permeability
antioxidant properties of MEL has been shown in patients with
transition and cytochrome c release.
rheumatoid arthritis (RA) with infertility in
It is of interest that MEL acts as an antiapoptotic agent in
elderly patients with primary essential hypertension How-
mitochondrial ROS/RNS- mediated cell death, but it also has
ever, there is no agreement about the beneﬁcial effects of MEL on
proapoptotic effects in several tumor cell lines such as in MCF-7
RA. Instead, MEL has been reported to be a RA promoter due to
breast cancer cells suggests that MEL has a potential
its capability to act as an immunoenhancing agent and stimula-
use to kill cancer cells preserving normal cells.
tor of proinﬂammatory cytokine release With regards to
A direct relationship between the indolamine and aging has
the enzymes of the antioxidant system, MEL regulates the expres-
not been proven but certain properties of the hormone indicate
sion of several genes such as those of superoxide dismutase (SOD)
its potential beneﬁt in the elderly. Data obtained in senescence-
and glutathione peroxidase (GPx). The hormone inﬂuences the
accelerated mice have demonstrated that chronic administration
enzyme activity and cellular mRNA levels of these proteins under
of MEL does not reduces respiratory function, decreases the neu-
physiological and oxidant conditions MEL concentration
ral and hepatic peroxidation products and the protein carbonyl
to up-regulate the enzyme gene expression corresponds to the
content in brain
physiological night time peak of plasma MEL, whereas higher con-
centration does not affect the gene expression. The antioxidant
effect of low concentrations of MEL seems to be indirect via up-
5. MEL and immune system
regulation of GPx gene expression and antioxidant activity, while
at high concentration the effect is attributed to direct radical scav-
A link between MEL from the pineal gland and the immune sys-
enging actions. It has been proposed that the ability of MEL to
tem in different species, including humans, has been documented
up-regulate the antioxidant enzyme involves both membrane and
immune system is modulated by different environmental
nuclear receptors appears that the stimulation of GPx by MEL
signals being light one of them. Although most of the light received
is the most consistent effect, while the effect on other enzymes is
by the retina goes to the visual cortex, another pathway from the
tissue-speciﬁc or conditional.
retina relays to the SCN, which forms part of the hypothalamic
A. Carpentieri et al. / Pharmacological Research 65 (2012) 437–444
region in the brain hypophysis and the pineal gland are
that MEL could be a mediator of acid-induced secretion
also involved in neuroendocrine changes induced by the light. MEL
of the MEL effects on the GI tract seem to be mediated by MT2
is one of the neuroendocrine hormones that are sensitive to changes
receptors the presence of MT1 receptor has been
in circadian rhythm. Since MEL is produced not only by the pineal
also demonstrated in human gallbladder epithelium from patients
gland, but also in the retina and in other parts of the body, the
with cholelithiasis and gallbladder carcinoma et al.
immune system might be affected by MEL originated in different
shown that MEL inhibits serotonin transporter activity in
organs. Besides, human peripheral blood mononuclear cells syn-
Caco-2 cells, apparently without mediation of MT1 or MT2 recep-
thesize important amounts of MEL, which indicates a potential
tors or PKC and cAMP pathways, but the doses used were high. It
intracrine and paracrine role of MEL in immune regulation. Mem-
is not known if this occurs under physiological conditions. MEL has
brane receptors have been found in T and B lymphocytes. MEL
also antioxidant properties in the GI system; its concentration in
also stimulates cytokine production by human peripheral blood
the human liver is suitable for prevention of oxidative damage. In
mononuclear cells through nuclear receptors Although the
our laboratory, MEL has been found to reverse the inhibition of the
mechanism by which MEL enhances the immune response is not
intestinal calcium absorption caused by menadione, apparently by
clear, it is thought that it may increase phagocytosis and antigen
antioxidant mechanisms
presentation. In addition, MEL induces T cell differentiation toward
MEL receptors have been detected in the pancreas and the
the type 1 helper T cells (Th1) phenotype and the activation of the
stimulation of pancreatic enzyme secretion by the hormone in a
immune system by the indolamine has been shown to be mediated
dose-dependent manner has been demonstrated. MEL was also
by the regulation of gene expression of cytokines in the spleen, thy-
implicated in the pancreatic endocrine function. Pinealectomy pro-
mus, lymph nodes and bone marrow killer cell activity
duces hyperinsulinemia and increases the triglyceride content in
and production are apparently increased by MEL administration
the liver of type 2 diabetic rats Endogenous MEL has been
either in humans or mice. Although many studies have implicated
shown to protect against pancreatitis, probably mediated by MT2
MEL as a positive regulator of immune system, other reports have
also suggested that MEL could act as an anti-inﬂammatory agent
through inhibition of immune responses. The anti-inﬂammatory
action is considered to be, at least in part, due to the induction
7. MEL and central nervous system
of type 2 helper T lymphocytes (Th2) producing interleukin (IL)-4
and inhibiting Th1 function has pleiotropic effects on dif-
MEL and melatoninergic drugs have hypnotic effects mediated
ferent steps of inﬂammation. It has a pro-inﬂammatory role at an
through MT1 and MT2 receptors, especially those in the SCN, which
early phase through activation of pro-inﬂammatory mediators such
acts on the hypothalamic sleep switch. They favor sleep initiation
as phosholipase A2 (PLA2), lipoxygenase (LOX) and cytokines such
and reset the circadian clock allowing persistent sleep, a require-
as IL-1 and tumor necrosis factor alpha (TNF␣). Contrarily, when
ment in circadian rhythm sleep alterations. The action of MEL on
the inﬂammatory process proceeds toward the chronic phase, MEL
sleep is mainly of a chronobiological nature. The hormone acts in
has a negative effect. The antagonist role of MEL on chronic inﬂam-
a dual way, resetting the clock via MT2 receptor and suppressing
mation is due to downregulation of mediators such as PLA2, LOX
neuronal ﬁring via MT1 receptor. Apparently, the soporiﬁc action
and cytokines, induction of a survival pathway in leukocytes and
of MEL also involves the thalamus. MEL receptors have been found
blocking of oxidation by its antioxidant properties
in this area and the formation of the spindles is stimulated by
MEL could also play a role in the inﬂuence of seasonal changes on
the hormone. Other areas of the brain seem to be involved in
the immune function. In healthy volunteers during autumn/winter
the transmission of MEL-dependent responses. It is uncertain to
season, the production of IL-6, interferon alpha (IFN-␣) and inter-
what extent the thalamus and other brain areas participate in the
feron gamma (IFN-␥) was higher. The seasonal changes in immune
sleep promotion as compared to the hypothalamic via MEL
functions appear to be mediated by alterations in duration of MEL
treatment to patients with severe CNS dysfunction and MEL deﬁ-
secretion. In this sense, changes in mood and behavior such as in
ciency is not sufﬁcient to mitigate sleep difﬁculties. In addition,
the seasonal affective disorder have been associated with seasonal
CNS destruction causing sleep fragmentation and loss of circadian
changes in cytokines like IL-6, IFN-␣ or the balance between Th1
rhythmicity, allows spindle formation. MEL has a different mode
and Th2 responses
of action as compared to other hypnotics such as benzodiazepins
and z-drugs, which produce a more generalized CNS depression
through GABA receptors. Only very high pharmacological doses
6. MEL and the gastrointestinal (GI) tract
of MEL can produce generalized sedative effects or even narcotic
effects but mediated by other mechanisms, such as antiexcita-
MEL is also produced in high amount in the enteroendocrine
tory suppression of calcium signaling and inhibition of neuronal
cells of GI mucosa. The amount of MEL in the gut is about 400 times
NO synthase. The major obstacle for the use of MEL in primary
larger than the content of MEL in the pineal gland pro-
chronic insomnia is that the half life of circulation is extremely
duction in the GI tract does not follow a circadian rhythm as occurs
short, 20–30 min and even less (eventually it could last up to a max-
in the pineal gland, but it responds to the periodicity of food intake
imum of 45 min). MEL promotes sleep initiation but improves sleep
rich in tryptophan a pinealectomy, serum levels of
maintenance only marginally. Part of this problem has been solved,
MEL do not maintain a rhythm according to the light–dark cycle,
at least in part, by drugs that prolong the release of MEL and the
but values are not too low at daytime because of the contribution
melatoninergic agonists with longer half-life
of MEL by the extra-pineal sources, mainly the GI tract
Neurodegenerative diseases such as Alzheimer disease (AD) and
The main enzymes of MEL synthesis, AANAT and HIOMT, have
Parkinson disease (PD) are age-related disorders that share mito-
also been detected in GI mucosa, supporting the hypothesis that in
chondrial dysfunction, oxidative/nitrosative stress and apoptosis
the gut MEL is synthesized from l-tryptophan present in food
in different areas of the brain. Patients with AD have a reduc-
The function of gut MEL is not clear yet because the intestine is
tion in MEL levels both in blood and cerebrospinal ﬂuid, which
not only source but also sink of MEL, brought by the circulation
is even present in preclinical stages MEL's cognitive bene-
Decreases in motility and increases in mucosal blood ﬂow
ﬁts have been demonstrated either in AD patients or in AD mice.
have been observed It has been also demonstrated that
Low levels of mRNA of SOD-1, GPx and catalase were found in hip-
luminal MEL stimulates duodenal HCO −
secretion, which suggests
pocampus from MEL treated AD mice In PD model animals,
A. Carpentieri et al. / Pharmacological Research 65 (2012) 437–444
MEL normalizes complex I activity from electron transport chain
various organs such as heart, kidney and bone marrow in the course
and oxidative status in mitochondria from substantia nigra and
of chemotherapy and radiotherapy. This protective effect of MEL
striatum and reduces mitochondrial inducible nitric oxide synthase
administration might allow treating patients more effectively using
decreasing nitric oxide radicals and preventing the inhibition of
higher doses of cytostatic drugs and radiation, which by themselves
complex IV by this radical treatment improved the quality
could damage the organs involved it has been shown
of sleep in patients with Parkinson idea that MEL is favor-
that MEL synergistically with vitamin D down-regulates Akt and
able in PD patients or PD animal models is controversial. Willis et al.
MDM2 leading to growth inhibition of breast cancer cells
that the light exposure at the bed time reduces the
In studies in which gastric damage was induced by
severity of PD symptoms, whereas the MEL intracerebroventricu-
indomethacin acetylsalicylic acid MEL prevented
lar implants increased its severity. Further studies about the use of
or reduced the gastric mucosal lesion. In addition, the combination
MEL in the PD treatment are needed.
of MEL with anti-ulcer drugs such as ranitidine and omeprazole
reduces the doses and minimizes the side effects
The pathophysiology of primary headache has been partially
8. MEL and cancer
attributed to desynchrony and dysfunction of the whole or part of
the retino-hypothalamic-pineal axis synchronizing ability
There is abundant evidence indicating that MEL is involved
of MEL together with its antioxidant and anti-inﬂammatory prop-
in preventing tumor initiation, promotion, and progression. The
erties, make this indolamine effective to decrease the frequency
increased incidence of breast, endometrial and colorectal cancer
or totally suppress headache of different kinds (migraine, cluster
seen in nurses and in other night shift workers suggests a possible
headache, etc.) allowing the patient to reduce the consumption of
link between diminished secretion of MEL and increased exposure
to light during nightime This evidence and the antioxidant
The use of MEL alone or in combination with other effective
and anti or proapoptotic action of MEL and its relationship with
agents has been reported to diminish collateral damage in sev-
the endocrine and immune systems led to examine the effect of
eral psychiatric disorders. Anxiolytic, sedative, anticonvulsant and
the hormone on tumor cells. Recent human and animal studies
anti-hypertensive effects of MEL may be helpful in the treatment
have shown that MEL also has important oncostatic properties by
of depression with inhibitors of monoaminooxidase. A combina-
altering the expression of anti-cancer cytokines IL-2 and IL-12 in
tion of MEL with lithium or valproate was more effective than
human neoplasms of 1 nM MEL to MCF-7 cell culture
either lithium or valproate alone in bipolar syndrome patients. MEL
inhibits proliferation the expression of proapoptotic
has also protective properties against haloperidol adverse effect in
proteins such as p53 and p21 reduces their metastatic
schizophrenia patients should be noted that many of the
potential due to increased expression of E-cadherin and ␤
beneﬁcial effects of MEL are associated with the improvement of
proteins antiproliferative effects on human breast cancer
sleep disorders that are very frequent in several neuropsychiatric
cells seem to be mediated by MT1 receptor, which suggests that
MEL or its analogs could be used as potential antitumoral agents
The administration of MEL alone is successful in the treatment
of circadian system disorders such as jet lag, shift work, some
The ﬁnding of MEL binding sites in human colon tissue suggested
sleeps disturbances, etc. In these cases MEL acts as a synchronizer,
a possible role of MEL in colorectal cancer. Schernhammer et al.
entraining the human activity circadian rhythms according to a
have found increased risk of colorectal cancer in nurses subjected to
phase-response curve
shift work, which was attributed to the suppression of MEL produc-
As shown in MEL is a pleiotropic molecule and its
tion by nocturnal lighting. Farriol et al. demonstrated that
administration to humans and animals at both physiological and
MEL was able to inhibit cell growth in CT-26 cells, a cell line derived
pharmacological concentrations seems to be non-toxic. Since MEL
from a murine colon carcinoma. Although MEL had no effect on cell
has a wide therapeutic range, it can be administered in differ-
growth at low doses, a signiﬁcant and progressive suppression of
ent doses according to the expected effects. The ability of MEL to
DNA synthesis was found with high doses of the indolamine. The
scavenge ROS is one of the explanations for the reduction of side
authors concluded that MEL exerted its anti-proliferative action
effects produced by some pharmacological agents such as onco-
through a non-hormone dependent mechanism, because no recep-
static, antinﬂammatory, antiepilepsic drugs these cases, the
tors were involved in the cell line selected. The oncostatic effect
doses of MEL range from 10 to 50 mg, much higher than physiolog-
of MEL on colon cancer was demonstrated to be mediated through
ical level, in order to counteract the high amounts of free radicals
MT2 receptors and through its binding to nuclear RZR/ROR ␣ recep-
Finally, an illustrative list of some recent in vivo
and in vitro studies exemplifying the effects of the combination
9. Combined effects of MEL with other drugs
of MEL with other drugs. As demonstrated, MEL can be useful in
the treatment of a wide variety of diseases improving their efﬁ-
The effect of MEL in the presence of other drugs has been
cacy while reducing the side effects in order to enhance life quality.
studied in different tissues. Several ﬁndings support the hypoth-
This broad therapeutic spectrum makes MEL a potential therapeu-
esis that MEL enhances the effect of chemotherapy on colorectal
tic tool in combination with other traditional therapies and opens
carcinoma. Cerea et al. the effect of the simultane-
a promising ﬁeld of research in the pharmacological area.
ous administration of MEL and the cytotoxic drug CPT-11 in 30
patients with metastatic colorectal carcinoma. It was found that
co-administration of MEL (20 mg/day at bedtime) with CPT-11 was
10. Remarks
more effective in controlling the disease than administration of
CPT-11 alone. It has also been shown that 1 mM MEL potentiates
MEL is a hormone produced in the pineal gland mainly at night
ﬂavone-induced apoptosis in HT-29 human colon cancer cells
time, but is also synthesized by other tissues being the intestine the
by increasing the level of oxidizable substrates that can be incor-
main source during the day. Its lipophilic nature makes MEL easily
porated into mitochondria in the presence of ﬂavones.
cross all membranes. Most of its actions are mediated by membrane
Another relevant aspect related to the application of MEL
and nuclear receptors. It has antioxidant and antiapoptotic prop-
as adjuvant in tumor therapy includes its protective effect on
erties, resulting in the improvement of mitochondrial metabolic